Mammalian cells respond to extracellular stimuli by activating signaling cascades that are mediated by members of the mitogen-activated protein (MAP) kinase family, which include the extracellular signal regulated kinases (ERKs), the p38 MAP kinases and the c-Jun N-terminal kinases (JNKs). MAP kinases are serine/threonine kinases that are activated by dual phosphorylation of threonine and tyrosine at the Thr-X-Tyr segment in the activation loop. MAP kinases phosphorylate various substrates including transcription factors, which in turn regulate the expression of specific sets of genes and thus mediate a specific response to the stimulus.
Three distinct genes, Jnk1, Jnk2, Jnk3 have been identified and at least ten different splicing isoforms of JNK exist in mammalian cells [S. Gupta et al., EMBO J., 15, pp. 2760-2770 (1996)]. Members of the JNK kinases are activated by proinflammatory cytokines tumor necrosis factor-alpha and interleukin-1 beta as well as environmental stress, such as anisomycin, UV irradiation, hypoxia, and osmotic shock [A. Minden et al., Biochemica et Biophysica Acta, 1333, F85-F104 (1997)]. Regulation & function of the JNK subgroup of MAP kinases. The down-stream substrates of JNKs include transcription factors c-Jun, ATF-2, Elk1, p53 and a cell death domain protein (DENN) [Y. Zhang et al. Proc. Natl. Acad. Sci. USA, 95, pp. 2586-2591 (1998)]. Each JNK isoform binds to these substrates with different affinities, suggesting a regulation of signaling pathways by substrate specificity of different JNKs in vivo (S. Gupta et al., 1996)
JNK1 and JNK2 are widely expressed in a variety of tissues. In contrast, JNK3 is selectively expressed in the brain and to a lesser extent in the heart and testis [S. Gupta et al., (1996); A. A. Mohit et al., Neuron, 14, pp. 67-78 (1995); J. H. Martin et al., Brain Res. Mol. Brain. Res., 35, pp. 47-57 (1996)]. In the adult human brain, JNK3 expression is localized to a subpopulation of pyramidal neurons in the CA1, CA4 and subiculum regions of the hippocampus and layers 3 and 5 of the neocortex [A. A. Mohit et al. (1995)]. The CA1 neurons of patients with acute hypoxia showed strong nuclear JNK3-immunoreactivity compared to minimal, diffuse cytoplasmic staining of the hippocampal neurons from brain tissues of normal patients [Y. Zhang et al. (1998)]. In addition, JNK3 co-localizes immunochemically with neurons vulnerable in Alzheimer's disease [A. A. Mohit et al., (1995)]. Disruption of the JNK3 gene caused resistance of mice to the excitotoxic glutamate receptor agonist kainic acid, including the effects on seizure activity, AP-1 transcriptional activity and apoptosis of hippocampal neurons, indicating that the JNK3 signaling pathway is a critical component in the pathogenesis of glutamate neurotoxicity (D. D. Yang et al., Nature, 389, pp. 865-870 (1997)]. Thus, selective modulation of JNK3 activity could potentially provide therapeutic intervention for neurodegenerative diseases such as stroke and epilepsy.
Despite the fact that the genes for various JNKs have been isolated and the amino acid sequences are known, no one has described X-ray crystal structural coordinate information of any of the JNKs. Such information would be extremely useful in identifying and designing potential inhibitors of various JNKs which, in turn, could have therapeutic utility.